Preparation of Mouse Brain Tissue for Immunoelectron Microscopy

Transmission electron microscopy (TEM) is extremely useful for visualizing microglial, oligodendrocytic, astrocytic, and neuronal subcellular compartments (dendrite, dendritic spine, axon, axon terminal, perikaryon), as well as their intracellular organelles and cytoskeleton, in the central nervous system at high spatial resolution. Combined with TEM, pre-embedding immunocytochemistry allows the discrimination of cellular elements with few distinctive features and identification criteria (e.g., microglial perikarya and processes, when using an antibody against the microglia-specific marker Iba1 (ionized calcium binding adaptor molecule 1; as presented here)), identifying the neurotransmitter contents of cellular elements (e.g., serotonergic) and their ultrastructural localization of soluble or membrane-bound proteins (e.g., 5 HT1A and EphA4 receptors). Here, we describe a protocol for transcardiac perfusion of mice with acrolein fixative, removal and sectioning of the brain, as well as immunoperoxidase-diaminobenzidine (DAB) staining, resin embedding, and ultrathin sectioning of the brain sections. Upon completion of these procedures, the immunostained material is ready for examination with TEM. When rigorously performed, this technique provides an excellent compromise between optimal ultrastructural preservation and immunocytochemical detection.Download video file.^{(101M, mp4)}     

Allosteric regulation of glycogen synthase controls glycogen synthesis in muscle

Glycogen synthase (GS), a key enzyme in glycogen synthesis, is activated by the allosteric stimulator glucose-6-phosphate (G6P) and by dephosphorylation through inactivation of GS kinase-3 with insulin. The relative importance of these two regulatory mechanisms in controlling GS is not established, mainly due to the complex interplay between multiple phosphorylation sites and allosteric effectors. Here we identify a residue that plays an important role in the allosteric activation of GS by G6P. We generated knockin mice in which wild-type muscle GS was replaced by a mutant that could not be activated by G6P but could still be activated normally by dephosphorylation. We demonstrate that knockin mice expressing the G6P-insensitive mutant display an ～80% reduced muscle glycogen synthesis by insulin and markedly reduced glycogen levels. Our study provides genetic evidence that allosteric activation of GS is the primary mechanism by which insulin promotes muscle glycogen accumulation in?vivo.     

Transcriptome Profile of a Long-Juvenile Soybean Genotype Huaxia-3 Under Short and Long Photoperiod

Plant Molecular Biology Reporter - The j allele delays flowering and enhances yield of long juvenile (LJ) soybean under short day (SD) condition. However, the underlying mechanism of j in flowering...     

Biomolecular phases in transverse palatal distraction: A review

Transverse palatal distraction is a biological process of regenerating new bone and enveloping soft tissues in the maxillary palate region. This technique is similar to Osteo-distraction (OD) procedure for bone lengthening in which gradual and controlled traction forces are applied on the osteotomy gaps to produce new bone in between the surgically separated bone segments. This review describes the different phases after osteotomy and the biological process involved during the new bone and soft tissue formation. The mechanical environment formed in the distraction area is due to the traction forces by the distractor appliance. This environment stimulates differentiation of pluripotent cells, neovascularization, osteogenesis and remodeling of newly formed bone. The role of different pro-inflammatory cytokines, interleukins, bone morphogenic proteins, transforming growth factors, fibroblast growth factors-2) and extracellular matrix proteins (osteonectin, osteopontin) during the distraction phases has been described in detail. Also, an important note on the nutritional aspect during Osteo-distraction will benefit the clinicians to guide their patients after osteotomy throughout the distraction process.     

The effects of a constant sprint-to-rest ratio and recovery mode on repeated sprint performance

It is unclear if a constant sprint-to-rest ratio allows full performance recovery between repeated sprints over different distances. This is important for the development of sprint-training programs. Additionally, there is conflicting evidence on whether active recovery enhances sprint performance. Three repeated sprint protocols were used (22 × 15, 13 × 30, and 8 × 50 m), with each having an active and passive recovery. Each trial was conducted with an initial sprint-to-rest ratio of 1:10. Repeated sprints were analyzed by comparing the first sprint to the last sprint. For the 15-m trials, there were no significant main effects for recovery or time and no significant interaction. For the 30-m trials, there was no main effect for recovery, but a main effect for time (F[1,10] = 15.995, p = 0.003; mean difference = 0.20 seconds, 95% confidence interval [CI] = 0.09-0.31 seconds, d = 1.4 [large effect]). There was no interaction of recovery and time in the 30-m trials. For the 50-m trials, there was no main effect for recovery, but a main effect for time (F[1,10] = 34.225, p = 0.0002; mean difference = 0.39 seconds, 95% CI = 0.24-0.55 seconds, d = 1.3 [large effect]). There was no interaction of recovery and time in the 50-m trials. The results demonstrate that a 1:10 sprint-to-rest ratio allows full performance recovery between 15-m sprints, but not between sprints of 30 or 50 m, and that recovery mode did not influence repeated sprint performance.     

The Effect of Chronic Long-Term Intermittent Hypobaric Hypoxia on Bone Mineral Density in Rats: Role of Nitric Oxide

Intermittent hypoxia is the most common pattern of hypoxic exposure in humans. The effect of chronic long-term intermittent hypobaric hypoxia (CLTIHH) on bone metabolism is not investigated. We examined the effect of CLTIHH on bone metabolism and the role of nitric oxide (NO) in this process. The rats were divided into three groups in this study. The animals in groups I and II have been exposed to CLTIHH. The animals in group II were also treated with nitric oxide synthase inhibitor NG-nitro-l-arginine methyl ester. To obtain CLTIHH, rats were placed in a hypobaric chamber (430 mm Hg; 5 h/day, 5 days/week, 5 weeks). The group III (control) rats breathed room air in the same environment. At the begining of the experiments, bone mineral density (BMD) of the animals were measured, and blood samples were collected from the tail vein. After the 5-week CLTIHH period, the same measurements were repeated. Parathyroid hormone, calcium, phosphate, bone alkaline phosphatase (b-ALP), NO, interleukin 1 beta, interleukin 6, and tumor necrosis factor alpha levels were determined. The cytokines, NO levels, and BMD in CLTIHH-induced rats were higher compared with baseline and control values. The cytokines, b-ALP, and BMD increased while NO levels decreased in the group II compared with baseline values. BMD values of group II were lower than group I but higher than control group. Our results suggested that CLTIHH has positive effects on bone density. Intermittent hypoxia protocols may be developed for treatment and prevention of osteopenia and osteoporosis.     

Castor and camphor essential oils alter hemocyte populations and induce biochemical changes in larvae of Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae)

Two plant essential oils; camphor and castor were tested for insecticidal and antifeedant activity against the 4th instar larvae of Spodoptera littoralis, a serious pest on cotton in Egypt. Also the impact of LC₁₀ of both oils on some physiological parameters in larvae was studied by using leaf dipping technique. Analysis of both oils using GC–MS revealed several insecticidal and antifeedant compounds. Our results showed higher insecticidal activity and antifeedant index of camphor oil against S. littoralis. The LC₅₀ and the antifeedant indices were 163.1, 246.8?mg/ml and 12.69, 6.62% for camphor and castor bean oil, respectively. The total hemocyte count (THC) and differential hemocyte count (DHC) were reduced significantly after 48?h of treatment compared to controls. Both oils reduced all types of hemocytes except plasmatocytes which were reduced only by castor oil. Camphor oil decreased total proteins and carbohydrates while castor oil targeted only carbohydrate content. Both oils didn't affect the amount of total lipids. Lipase, α-amylase and glucose-6-phosphate dehydrogenase (G6PD) enzyme activities were increased significantly in larvae treated with camphor oil than other treatments. These results clearly indicate that castor and camphor oils can affect the nutritional status in S. littoralis larvae, thereby changing the internal metabolic processes in the larvae which make them as potential control agents in IPM programs against S. littoralis.